Pyrimidine intermediates

ABSTRACT

The present invention is directed to compounds of formula (I), ##STR1## wherein R 1  is OH, NH 2  ; R 2  is a heteroaromatic or aromatic substituent; R 3  is H, OH, F, OCH 3  ; R 4  is H, F, OH or an ether or ester residue thereof, OCH 3 , CN, C═CH, N 3  ; R 5  is OH or an ether or ester residue thereof including mono, di- and triphosphate esters (α), wherein n is 0 or 1 and M is hydrogen or a pharmaceutically acceptable counterion such as sodium, potassium, ammonium or alkylammonium; and pharmaceutically acceptable salts thereof; and pharmaceutical compositions comprising said compounds can be used for therapeutic treatment of virus infections. The present invention is also directed to compounds of formula (I&#39;), ##STR2## wherein R 1  and R 2  are as defined above, as intermediates.

FIELD OF THE INVENTION

The present invention relates to novel chemical compounds andpharmaceutically acceptable salts thereof which can be used in therapyfor therapeutic treatment of the acquired immuno deficiency syndrome(AIDS) and infections caused by viruses requiring reverse transcriptasefor replication, such as human immuno deficiency virus and hepatitis Bvirus, and also for treatment of virus diseases, such as those of herpesviruses diseases which include common infections and neoplasticdiseases, i.e. cancer. The invention also relates to novel precursorcompounds constituting a further aspect of the invention.

BACKGROUND OF THE INVENTION

The effects of viruses on bodily functions is the end result of changesoccurring at the cellular and subcellular levels. The pathogenic changesat the cellular level are different for different combinations ofviruses and host cells. While some viruses cause a general destruction(killing) of certain cells, other may transform cells into a neoplasticstate.

Important common viral infections are herpes dermatitis (includingherpes labialis), herpes keratitis, herpes genitalis, herpes zoster,herpes encephalitis, infectious mononucleosis and cytomegalovirusinfections all of which are caused by viruses belonging to the herpesvirus group. Other important viral diseases are influenza A and B whichare caused by influenza A and B virus respectively. Another importantcommon viral disease is viral hepatitis and especially hepatitis B virusinfections are widely spread. Effective and selective antiviral agentsare needed for treatment of these diseases as well as for other diseasescaused by viruses. Several different viruses of both DNA and RNA typehave been shown to cause tumors in animals. The effect of cancerogenicchemicals can on animals result in activation of latent tumor viruses.It is possible that tumor viruses are involved in human tumors. The mostlikely human cases known today are leukemias, sarcomas, breastcarcinomas, Burkitt lymphomas, nasopharyngeal carcinomas and cervicalcancers where RNA tumor viruses and herpes viruses are indicted andpapillomas where papilloma viruses are involved. This makes the searchfor selective inhibitors of tumorogenic viruses and their functions animportant undertaking in the efforts to treat cancer.

In the late seventies a new disease was reported, which subsequently wasreferred to as Acquired Immuno Deficiency Syndrome (AIDS). It is nowgenerally accepted that a retrovirus referred to as HIV (HumanImmunodeficiency Virus), formerly known as Human T-cell LymphotropicVirus (HTLV-III) or Lymphadenopathy Associated Virus (LAV) plays anessential role in the etiology of AIDS. Different types of HIV have beenfound, such as HIV-1 and HIV-2 and more are likely to be isolated.

AIDS is characterized by a profound immunodeficiency due to low numbersof a subset of lymphocyte-T-helper cells, which are one target for HIVinfection. The profound immunodeficiency in AIDS patients makes thesepatients highly susceptible to a variety of opportunistic infections ofbacterial, fungal, protozoal or viral etiology. The etiological agentsamong viral oppportunistic infections are often found in the herpesvirus group, i.e. herpes simplex virus (HSV), Varicella Zoster virus(VZV), Epstein-Barr virus (EBV) and, especially, cytomegalovirus (CMV).Other retroviruses affecting humans are HTLV-I and II and examples ofretroviruses affecting animals are feline leukemia virus and equineinfectious anaemia virus. Human diseases such as multiple sclerosis,psoriasis, tropical spastic paresis and Kawasaki disease have also beenreported to be associated with retrovirus infections. Hepatitis B virusinfections cause severe disease such as acute hepatitis, chronichepatitis, fulminant hepatitis in a considerable number of persons. Itis estimated that there are 200 million patients with chronic hepatitisB infection in the world. A considerable number of the chronic casesprogress to liver cirrosis and liver tumours. In some cases thehepatitis infections also take a rapid and severe course as in fulminantB hepatitis with about 90% mortality. At present there is no knowneffective treatment against hepatitis B infections. The replication ofhepatitis B virus is similar to that of retroviruses and it contains thesame essential viral reverse transcriptase activity.

GENERAL OUTLINE OF THE INVENTION

A great number of nucleoside analogues exhibit several antimetabolicactivities. They do so by substituting for or competing with thenaturally occuring nucleosides. Recently, some nucleoside analogues havebeen described, which inhibit in cell culture the multiplication ofhuman immunodeficiency virus (HIV, also called HTLV-III, LAV) thecausative agent of AIDS and AIDS-related complex (ARC).

WE have now found that activities for inhibition of herpesmultiplication are exhibited by nucleoside analogues, in which thepyrimidine bases are substituted in the 5-position by a heteroaromatic,or aromatic substituent. The nucleoside analogues may be either alpha-or beta-anomers.

DISCLOSURE OF THE INVENTION

The present invention relates to new compounds of the formula I ##STR3##wherein the radicals R¹, R², R³, R⁴ and R⁵ are defined as follows:

R¹ : OH, NH₂ ;

R² : ##STR4## wherein X is O, S, N--R⁷, Se;

R⁶ is H, straight or branched C₁₋₁₀ alkyl, F, Cl, Br, I,

X--R⁷, --CH═CH--R⁷, --C≡C--R⁷, CO₂ R⁷, CH₂ X--R⁷ ;

R⁷ is H, straight or branched C₁₋₅ alkyl, phenyl;

R³ : H, OH, F, OCH₃ ;

R⁴ : H, F, OH or an ether or ester residue thereof, OCH₃, CN, C≡CH, N₃ ;

R⁵ : OH or an ether or ester residue thereof; ##STR5## wherein n is 0 or1 and M is hydrogen or a pharmaceutically acceptable counterion such assodium, potassium, ammonium or alkylammonium; and pharmaceuticallyacceptable salts thereof.

The invention consequently also refers to the compounds of the formula Ifor use in therapy. The compounds of the formula I are useful astherapeutic agents in the control and treatment of virus infections inman. The compounds of the formula I are useful as therapeutic agents inthe control and treatment of infections caused by hepatitis B virus inmammals and man.

All retroviruses, including HIV, require the enzyme reversetranscriptase in their natural cycle of replication.

Hepatitis B virus (HBV) is a DNA virus with a unique circulardouble-stranded DNA genome which is partly single-stranded. It containsa specific DNA polymerase required for viral replication. This DNApolymerase also acts as a reverse transcriptase during the replicationof HBV DNA via an RNA intermediate.

The compounds of the formula I inhibit the activity of DNA polymerase ofhepatitis B virus.

Another important area of use for the compounds of the formula I is inthe treatment of herpes virus infections. Among the herpes viruses maybe mentioned Herpes simplex type 1 and 2, vericella (Herpes zoster),virus causing infectious mononucleosis (i.e. Epstein-Barr virus),cytomegalovirus and human herpes virus type 6. Important diseases causedby herpes viruses are herpes dermatitis (including herpes labialis),herpes genitalis, herpes keratitis, herpes encephalitis and herpeszoster.

The invention further provides:

A pharmaceutical composition comprising a compound of the formula I asan active ingredient and a pharmaceutically acceptable carrier,including lipsomes; and

A method for therapeutic treatment of virus infections in an animal orhuman host in need of treatment comprising administering an effectiveamount of a compound of the formula I.

It is a preferred aspect of the invention to treat infections caused byherpes viruses and hepatitis B virus.

The nucleoside analogues of the invention are composed of a5-substituted uracil or cytosine base and a sugar moiety which can forinstance be ribose, 2'-deoxyribose, 2',3'-dideoxyribose, arabinose, oranalogues thereof.

Preferred compounds of the formula I ##STR6## are those wherein

R² is 2-furyl, 2-thienyl, selenienyl, thiazolyl, 2-(1-alkyl)pyrrolyl ormethoxyphenyl;

R³ is hydrogen, hydroxy or fluoro;

R⁴ is hydrogen, hydroxy, fluoro, cyano or azido; and

R⁵ is hydroxy, a mono-, di- or triphosphate thereof or ##STR7## whereinM is a pharmaceutically acceptable counterion.

Examples of especially preferred compounds are those of the formula Iwherein

    ______________________________________                                        R.sup.1 R.sup.2    R.sup.3                                                                              R.sup.4                                                                            R.sup.5                                        ______________________________________                                        OH      2-furyl    H      OH   OH or triphosphate                             OH      2-thienyl  H      OH   OH or triphosphate                             OH      2-selenienyl                                                                             H      OH   OH or triphosphate                             OH      2-thiazolyl                                                                              H      OH   OH or triphosphate                             OH      2-furyl    OH     OH   OH or triphosphate                             OH      2-thienyl  OH     OH   OH or triphosphate                             OH      2-selenienyl                                                                             OH     OH   OH or triphosphate                             OH      2-thiazolyl                                                                              OH     OH   OH or triphosphate                             OH      2-furyl    OH     F    OH or triphosphate                             OH      2-thienyl  OH     F    OH or triphosphate                             OH      2-selenienyl                                                                             OH     F    OH or triphosphate                             OH      2-thiazolyl                                                                              OH     F    OH or triphosphate                             OH      2-furyl    OH     N.sub.3                                                                            OH or triphosphate                             OH      2-thienyl  OH     N.sub.3                                                                            OH or triphosphate                             OH      2-selenienyl                                                                             OH     N.sub.3                                                                            OH or triphosphate                             OH      2-thiazolyl                                                                              OH     N.sub.3                                                                            OH or triphosphate                             OH      2-furyl    H      H    OH or triphosphate                             OH      2-thienyl  H      H    OH or triphosphate                             OH      2-selenienyl                                                                             H      H    OH or triphosphate                             OH      2-thiazolyl                                                                              H      H    OH or triphosphate                             OH      2-furyl    H      F    OH or triphosphate                             OH      2-thienyl  H      F    OH or triphosphate                             OH      2-selenienyl                                                                             H      F    OH or triphosphate                             OH      2-thiazolyl                                                                              H      F    OH or triphosphate                             OH      2-furyl    H      N.sub.3                                                                            OH or triphosphate                             OH      2-thienyl  H      N.sub.3                                                                            OH or triphosphate                             OH      2-selenienyl                                                                             H      N.sub.3                                                                            OH or triphosphate                             OH      2-thiazolyl                                                                              H      N.sub.3                                                                            OH or triphosphate                             OH      2-furyl    H      OH   methylphosphonate                              OH      2-thienyl  H      OH   methylphosphonate                              OH      2-thiazolyl                                                                              H      OH   methylphosphonate                              OH      2-furyl    H      H    methylphosphonate                              OH      2-thienyl  H      H    methylphosphonate                              OH      2-thiazolyl                                                                              H      H    methylphosphonate                              OH      2-furyl    H      F    methylphosphonate                              OH      2-thienyl  H      F    methylphosphonate                              OH      2-thiazolyl                                                                              H      F    methylphosphonate                              OH      2-furyl    H      N.sub.3                                                                            methylphosphonate                              OH      2-thienyl  H      N.sub.3                                                                            methylphosphonate                              OH      2-thiazolyl                                                                              H      N.sub.3                                                                            methylphosphonate                              OH      2-furyl    OH     F    methylphosphonate                              OH      2-thienyl  OH     F    methylphosphonate                              OH      2-thiazolyl                                                                              OH     F    methylphosphonate                              ______________________________________                                    

Esters and ethers of the nucleosides are also included in the invention.Examples of esters are mono-, di- and tri-phosphate esters, carboxylicesters, carbonate esters, carbamate esters and sulphonic esters. Theacid part of the esters may have alkyl, aryl or arylalkyl chains, wherethe aryl functionalities are optionally substituted for example byalkoxy, amino, nitrile, alkyl or sulphonamido groups or by one or morehalogen atoms. Examples of other types of derivatives of the nucleosidesare alkyl or arylalkyl derivatives of the 5-hydroxyl group. Thearylalkyl ether derivatives may be for example benzyl or tri-phenylmethyl and the aryl moiety may be optionally substituted. Furthermore,it is understood that the examples of the pharmaceutically acceptablesalts cited below also apply to the various esters of derivatives of thenucleosides of the invention.

In a compound of the formula I R⁵ as an ether residue can be defined asOR⁸, wherein R⁸ is C₁₋₆ alkyl, arylakyl optionally substituted with oneor more alkoxy, amino, nitrile or sulphamido groups or one or morehalogen atoms.

R⁴ and R⁵ as an ester residue can be derived from a carboxylic acid R⁹COOH, a carbonic acid R¹⁰ OCOOH, a double ester of a carbonic acid R¹⁰CO₂ CH(R¹¹)OCO₂ H, a sulphonic acid R¹⁰ SO₂ OH, a carbamic acid R¹⁰NHCOOH or a phosphoric acid, wherein R⁹ is hydrogen, C₁₋₁₇ alkyl,alkoxyalkyl, arylalkyl or aryl, R¹⁰ is C₁₋₁₇, arylalkyl or aryl, R¹¹ ishydrogen or C₁₋₃ alkyl and said aryl and arylalkyl groups optionally canbe substituted with one or more alkyl, alkoxy, amino, nitrilesulphonamido groups or one or more halogen atoms.

Examples of pharmaceutically acceptable salts of the compounds offormula I include base salts, e.g. derived from an appropriate base,such as alkali metal (e.g. sodium, potassium, alkaline earth metal, e.g.magnesium) salts, ammonium and NX₄ ⁺ (wherein X is C₁₋₄ alkyl).Physiologically acceptable acid salts include salts of organiccarboxylic acids such as acetic, lactic, gluconic, citric, tartaric,maleic, malic, pantothenic, isethionic oxalic, lactobionic and succinicacids; organic sulfonic acids such as methanesulfonic, ethanesulfonic,benzenesulfonic, p-chlorobenzenesulphonic and p-toluenesulfonic acidsand inorganic acids such as hydrochloric, hydroiodic, sulfuric,phosphoric and sulfamic acids.

Physiologically acceptable counterions M of the phosphonate groupsinclude inorganic and organic counterions. Inorganic counterions are forexample ammonium, sodium, potassium, lithium, magnesium and calcium.Organic counterions are derived from non-toxic bases, such as primary,secondary and tertiary amines, including naturally occuring amines.Examples of such amines are diethylamine, triethylamine, isopropylamine,ethanolamine, morpholine, 2-diethylaminoethanol, glucosamine,N-methylglucamine, piperazine and dicyclohexylamine.

In clinical practice the pyrimidine derivatives of the formula I willnormally be administered orally, by injection or by infusion in the formof a pharmaceutical preparation comprising the active ingredient in theform of the original compound or optionally in the form of apharmaceutically acceptable salt thereof, in association with apharmaceutically acceptable carrier which may be a solid, semi-solid orliquid diluent or an ingestible capsule. The compound may also be usedwithout carrier material. As examples of pharmaceutical preparations maybe mentioned tablets, dragees, capsules, granulates, suspensions,elixirs, syrups, solutions, liposomes etc. Usually the active substancewill comprise between 0.05 and 20% for preparations intended forinjection and between 10 and 90% for preparations intended for oraladministration.

In the treatment of patients suffering from hepatitis B virusinfections, it will be preferred to administer the compounds by anysuitable route including the oral, parenteral, rectal, nasal, topicaland vaginal route. The parenteral route includes subcutaneous,intramuscular, intravenous and sublingual administration. The topicalroute includes buccal and sublingual administration. The dosage at whichthe active ingredients are administered may vary within a wide range andwill depend on various factors such as the severity of the infection,the age of the patient etc., and may have to be individually adjusted.As a possible range for the amount of the compounds of the invention ora physiologically acceptable salt thereof to be administered per day maybe mentioned from about 10 mg to about 10,000 mg, preferentially 100-500mg for intravenous administration and preferentially 100-3000 mg fororal administration.

METHODS OF PREPARATION

The compounds of the invention may be prepared by one of the followinggeneral methods, constituting a further aspect of the invention.

A. Condensing a glycoside as comprised in formula I where the hydroxylgroups may be optionally protected to the N-1 position of a pyrimidinederivative, according to known methods described in the literature. Suchmethods are described for example in "Basic Principles in Nucleic AcidChemistry", Vol. 1 (Academic Press, 1974, Ed. P. O. P. Ts'o), in"Nucleoside Analogues, Chemistry, Biology and Medical Applications"(Pharma Press, 1979, Eds. R. T. Walker, E. De Clercq and F. Eckstein).##STR8##

Examples of suitable derivatives of the reacting species are thosewherein Z is Cl, Br, I, acyloxy or alkoxy; R'is an alkyl or silylprotecting group, such as C₂ H₅ or (CH₃)₃ Si; R^(1') is R¹ as definedabove, OC₂ H₅, (CH₃)₃ SiO, or N(COCH₃)Si(CH₃)₂ ; R² is as defined above;R^(3') and R^(4') is R³ and R⁴ respectively as defined above with theproviso that when R³ or R⁴ is OH said OH must be protected as O-acyl,O-benzoyl, O-benzyl or O-silyl (e.g. dimethyl, tert-butylsilyl); andR^(5') is R⁵ as defined above or OR⁸ wherein R⁸ is as defined above orsilyl (e.g. dimethyl, tert-butylsilyl). After condensation the productsmay be hydrolyzed or converted by conventional methods, known to thoseskilled in the art, into compounds of the formula I.

The glycosides are known or may be prepared by suitable adaptions ofknown methods. The syntheses of a2,3-dideoxy-3-fluoro-erythro-pentofuranoside for example, has beendescribed by G. W. J. Fleet and J. C. Son in Tetrahedron Letters 40(1987) pp 3615-3618. The other 3- substitutents may be introduced bymethods analogous to those described above and described by N. B.Dyathina and A. V. Azhayev in Syntheses 1984 pp 961-963. Thearabinosylglycosides may be prepared by similar methods.

B. The β-anomers of the arabinosyl-pyrimidine nucleoside analogues maybe prepared by hydrolysis of the corresponding 2,2'-anhydro nucleosideanalogues. ##STR9## wherein R^(1") is O or NH and R¹, R², R^(4') andR^(5') are as defined above. The hydrolysis may be performed byconventional methods, described in the literature and known to thoseskilled in the art. It may for example be performed by treating the2,2'-anhydronucleosides with an aqueous acid.

C. The halogeno, OCH₃, N₃, CN and C≡CH substituents in the 3'-positionof the glycon moiety may be introduced by substitution or a hydroxylgroup or a suitably derivatized hydroxyl group ##STR10## wherein Y is OHor a functionality that will be leaving in the substitution reactionsuch as for example CF₃ SO₃ ; and R^(1'), R², R^(3'), R⁴ and R^(5') areas defined above.

The following examples will further illustrate the invention:

Example 11-(2-Deoxy-3,5-di-O-p-toluoyl-alpha-D-ribofuranosyl)-5-(furyl)uracil(VSB 005) and Example 21-(2-Deoxy-3,5-di-O-p-toluoyl-beta-D-ribofuranosyl)-5-(furyl)uracil (VSB006)

5-(2-Furyl)uracil (150 mg, 0.84 mmol) in hexamethyldisilazane (10 ml)was heated at reflux for 5 hours together with chlorotrimethylsilane (10drops) and ammoniumsulfate (a few mg). The solution was filtered andevaporated in vacuo to dryness to give bis-trimethylsilylated5-(2-furyl)uracil (240 mg) as a crude product. This crude product wasdissolved in acetonitrile (15 ml, dried over molecular sieves) and addedto a solution of 2-deoxy-3,5-di-O-p-toluoyl)-D-arythro-pentosyl chloride(331 mg, 0.85 mmoles; prepared according to C. C. Bhat in SyntheticProcedures in Nucleic Acid Chemistry, Vol. 1, p. 521, Interscience Publ.1968; W. W. Zorbach and R. S. Tipson eds.) in dried acetonitrile (20 ml)and stirred over night at ambient temperature under an athmosphere ofnitrogen. The solution was filtered, evaporated in vacuo and the residuewas separated by chromatography on a column of silica to give puresamples of the alpha-anomer (62 mg) and of the beta-anomer (29 mg, m.p.190°-192° C.). Thin layer chromatography (silica,dichloromethane-ethylacetate 5-1) R_(f) : alpha 0.37; beta 0.50.

Example 31-(2-Deoxy-3,5-di-O-p-toluoyl-alpha-D-ribofuranosyl)-5-(2-thienyl)uracil(VSB 128) and Example 41-(2-deoxy-3,5-di-O-p-toluoyl-beta-D-ribofuranosyl)-5-(2-thienyl)uracil(VSB 125)

5-(2-Thienyl)uracil (0.97 g, 5 mmol) in hexamethyldisilazane (10 ml) washeated at reflux for about 2.5 hours together with chlorotrimethylsilane (10 drops) and ammoniumsulfate (a few mg). The solutionwas filtered and evaporated in vacuo to dryness to givebis-trimethylsilylated 5-(2-thienyl)uracil, which was dissolved in1,2-dichloroethane (25 ml, dried over molecular sieves) and added to asolution of 2-deoxy-3,5-di-O-p-toluoyl)-D-erythro-pentosyl chloride(1.55 g, 4 mmoles; prepared according to C. C. Bhat in SyntheticProcedures in Nucleic Acid Chemistry, Vol. 1, p. 521, Interscience Publ.1968; W. W. Zorbach and R. S. Tipson eds.) in dry, 1,2-dichloroethane(25 ml). Molecular sieves (2 g, 4 Å) was added and the mixture wasstirred at ambient temperature over night after which it was filtered.The solution was washed with an aqueous, saturated solution of sodiumbicarbonate (50 ml) and water (50 ml), dried over sodium sulfate,concentrated to a volume of about 25 ml and refrigerated. Theprecipitate was filtered and recrystallized from 1,2-dichloroethane togive pure β-anomer (0.70 g). The remaining combined solutions wereevaporated and the residue was separated on a column of silica elutedwith chloroform-ethyl acetate 5-1, to give the pure alpha-anomer, VSA128, (0.51 g, m.p. 201°-3° C.) and the pure beta-anomer, VSA 125, (totalcombined yield 0.86 g, m.p. 217°-9° C.). Thin layer chromatography(silica, chloroform-ethyl acetate 5-1) R_(f) : alpha 0.23; beta 0.30.

Analysis for C₂₉ H₂₆ N₂ O₇ S; calculated (found) %: alpha: C 63.72(63.5); H 4.80 (4.8); N 5.13 (5.0); beta: C63.72 (63.2); H 4.80 (4.8); N5.13 (5.1).

Analogous to examples 1 and 2, table 1 lists some further examples whichare characterized as shown in table 2.

                  TABLE 1                                                         ______________________________________                                        Examples of 1-(2-deoxy-3,5-di-O-p-toluoyl-alpha/beta-                         D-ribofuranosyl)-5-R.sup.2 -uracil compounds                                  Example alpha/beta R.sup.2                                                    ______________________________________                                         1      alpha      2-furyl                                                     2      beta       2-furyl                                                     3      alpha      2-thienyl                                                   4      beta       2-thienyl                                                   5      alpha      3-furyl                                                     6      alpha      3-thienyl                                                   7      beta       3-thienyl                                                   8      alpha      2-selenienyl                                                9      beta       2-selenienyl                                               10      alpha      3-selenienyl                                               11      beta       3-selenienyl                                               12      alpha      2-pyridyl                                                  13      alpha      3-pyridyl                                                  14      alpha      4-pyridyl                                                  15      alpha      2-(5-methyl) thienyl                                       16      beta       2-(5-methyl)thienyl                                        17      alpha      2-(5-hexyl)thienyl                                         18      beta       2-(5-hexyl)thienyl                                          55 56   alpha beta                                                                               ##STR11##                                                  57 58   alpha beta                                                                               ##STR12##                                                 59      alpha      2-methoxyphenyl                                            60      beta       2-methoxyphenyl                                            61      alpha      3-methoxyphenyl                                            ______________________________________                                    

                                      TABLE 2                                     __________________________________________________________________________    Data for 1-(2-deoxy-3,5-di-O-P-toluoyl-alpha/beta-D-ribofuranosyl)-5-R.sup    .2 -uracil compounds                                                          .sup.13 C NMR (CDCl.sub.3)δ                                                                 .sup.1 H NMR (CDCl.sub.3)δ                                                        m.p. Thin layer                                 Example                                                                            1' 2' 3' 4' 5' 1'        °C.                                                                         chromatography R.sub.f                     __________________________________________________________________________     1   88.1                                                                             39.4                                                                             74.9                                                                             85.8                                                                             64.3                                                                             6.44 d         0.37 a                                      2   85.8                                                                             38.7                                                                             75.1                                                                             83.3                                                                             64.7                                                                             6.53 t         0.50 a                                      3   88.0                                                                             39.2                                                                             74.6                                                                             85.7                                                                             64.0                                                                             6.41 d    201-3                                                                              0.23 b                                      4   85.9                                                                             38.8                                                                             75.0                                                                             83.5                                                                             64.4                                                                             6.48 t    217-9                                                                              0.30 b                                      5   88.1                                                                             39.3                                                                             74.8                                                                             85.8                                                                             64.2                                                                             6.33 d (J 3.5 Hz)                                                                       179-81                                                                             0.20 b                                      6                                                                             7                                                                             8                            182-4                                                                              0.22 b                                      9                            214-6                                                                              0.33 b                                     10                                                                            11                                                                            12   88.1                                                                             39.5                                                                             74.5                                                                             85.5                                                                             64.0                                                                             6.11 d (J 3.1 Hz)                                                                            0.12 a                                     13   88.1                                                                             39.2                                                                             74.8                                                                             85.8                                                                             64.1                                                                             6.41 d                                                    14                  6.44           0.10 c                                     15   88.0                                                                             39.2                                                                             74.8                                                                             85.7                                                                             64.2                                                                             6.41 d (J 3.3 Hz)                                                                            0.17 d                                     16   85.7                                                                             38.6                                                                             75.0                                                                             83.2                                                                             64.4                                                                             6.46 t         0.26 d                                     17   88.0                                                                             39.3                                                                             74.8                                                                             85.8                                                                             64.2                                                                             6.40 d (J 2.8 Hz)                                                                            0.22 d                                     18   85.7                                                                             38.5                                                                             75.0                                                                             83.2                                                                             64.4                                                                             6.49 t (J 2.0 Hz)                                                                            0.34 d                                     55                             98-101                                                                            0.57 a                                     56                            213-217                                                                            0.70 a                                     57                            111-115                                                                            0.41 a                                     58                            215-218                                                                            0.54 a                                     59                            88-90                                                                              0.23 a                                     60                            196-198                                                                            0.55 a                                     61                            167-169                                                                            0.28 a                                     __________________________________________________________________________     a) CH.sub.2 Cl.sub.2 --EtOAc 51; b) CHCl.sub.3 --EtOAc 51; c) CHCl.sub.3      --EtOAc 41; d) CHCl.sub.3 --EtOAc 91.                                    

Example 19 1-(2-Deoxy-alpha-D-ribofuranosyl)-5-(2-furyl)uracil (VSB 007)

VSB 005 (62 mg, 0.117 mmol) was suspended in methanol (15 ml, dried overmolecular sieves) and sodium methoxide in methanol (1.2 ml, 0.2M) wasadded. The mixture was stirred at ambient temperature under anathmosphere of nitrogen for 24 hours, after which an ion exchanger,Dowex 50 W×8 H⁺, was added. The solution was filtered and the solventevaporated in vacuo. The residue was purified on a column of silicaeluted with ethyl acetate-ethanol 9-1, to give1-(2-deoxy-α-D-ribofuranosyl)-5-(2-furyl)uracil 32 mg (93%). Thin layerchromatography (silica, ethyl acetate-ethanol 18-1) R_(f) : 0.42.

Example 20 1-(2-Deoxy-beta-D-ribofuranosyl)-5-(2-furyl)uracil VSB 008

VSB 006 (29 mg, 0.55 mmol) was hydrolyzed with sodium methoxide asdescribed for VSB 005. After completion of reaction, the dry residue ofthe crude product was triturated with hexane and purified on silica togive 1-(2-deoxy-β-D-ribofuranosyl)-5-(2-furyl)uracil. The thin layerchromatography (silica, ethyl acetate-ethanol 18-1) R_(f) : 0.47.

Example 21 1-(2-Deoxy-alpha-D-ribofuranosyl)-5-(2-thienyl)uracil (VSB134)

VSA 128 (0.35 g, 0.64 mmol) was dissolved in methanol (50 ml) and sodiummethoxide in methanol (5 ml, 0.2M) was added. The solution was stirredat ambient temperature over night, after which it was neutralized withDowex 50W×8 H⁺. The solution was filtered, evaporated in vacuo and theresidue was triturated with diethyl ether to give as a solid residue1-(2-deoxy-α-D-ribofuranosyl)-5-(2-thienyl)uracil. Thin layerchromatography (silica, chloroform-methanol 85-15) R_(f) : 0.44.Analysis for C₁₃ H₁₄ N₂ O₅ S, calculated (found) %: C 50.31 (50.3); H4.55 (4.5); N 9.03 (8.8).

Example 22 1-(2-Deoxy-beta-D-ribofuranosyl)-5-(2-thienyl)uracil (VSA133)

The title compound was prepared from VSA 125 (0.55 g, 1 mmol) in thesame way as has been described for the corresponding alpha-anomer VSA134. Thin layer chromatography for VSA 133 (silica, chloroform-methanol85-15) R_(f) : 0.47.

Analogous to examples 19-22, table 3 lists some further examples whichwere characterized as shown in table 4.

                  TABLE 3                                                         ______________________________________                                        Examples of 1-(2-deoxy-alpha/beta-                                            D-ribofuranosyl)-5-R.sup.2 uracil compounds                                   Example alpha/beta R.sup.2                                                    ______________________________________                                        19      alpha      2-furyl                                                    20      beta       2-furyl                                                    21      alpha      2-thienyl                                                  22      beta       2-thienyl                                                  23      alpha      3-furyl                                                    24      alpha      3-thienyl                                                  25      beta       3-thienyl                                                  26      alpha      2-selenienyl                                               27      beta       2-selenienyl                                               28      alpha      3-selenienyl                                               29      alpha      2-pyridyl                                                  30      alpha      3-pyridyl                                                  31      alpha      4-pyridyl                                                  32      alpha      2-(5-methyl)thienyl                                        33      beta       2-(5-methyl)thienyl                                        34      alpha      2-(5-hexyl)thienyl                                         35      beta       2-(5-hexyl)thienyl                                          62 63   alpha beta                                                                               ##STR13##                                                  64 65   alpha beta                                                                               ##STR14##                                                 ______________________________________                                    

                                      TABLE 4                                     __________________________________________________________________________    Data for 1-(2-deoxy-alpha/beta-D-ribofuranosyl)-5-R.sup.2 -uracil             compounds                                                                     __________________________________________________________________________    .sup.3 C NMR δ .sup.1 NMR δ                                                                        Thin layer                                                                              m.p.                               Example                                                                            1' 2' 3' 4' 5'  1'          chromatography R.sub.f                                                                  °C.                         __________________________________________________________________________    19   91.9                                                                             42.2                                                                             72.9                                                                             89.2                                                                             63.8 a                                                                            6.35 dd (J 2.9; 1 Hz)a                                                                    0.42 c                                       20   89.4                                                                             42.0                                                                             72.5                                                                             87.3                                                                             63.1 a                                                                            6.44 t (J 3.3 Hz)a                                                                        0.47 c                                       21   92.1                                                                             42.1                                                                             73.0                                                                             89.0                                                                             63.9 a                                                                            6.24 dd (J 2.9; 1.0 Hz)b                                                                  0.44 d                                       22   87.8                                                                             40.6                                                                             70.3                                                                             85.1                                                                             61.2 b                                                                            6.23 t (J 3.3 Hz)b                                                                        0.47 d                                       23   90.2                                                                             41.7                                                                             72.8                                                                             88.3                                                                             63.6 a                                                                            6.37 dd (J 3; 1 Hz)a                                                                      0.44 e                                       24                                                                            25                   6.36 t (J 5 Hz) a                                        26                               0.23 f                                       27                               0.29 f                                       28                   6.29 dd (J 5; 1 Hz)a                                     29   91.7                                                                             42.2                                                                             72.8                                                                             89.5                                                                             63.7 a                                                                            6.33 dd (J 1.1; 1.5 Hz)a                                                                  0.43 g                                       30   92.1                                                                             42.1                                                                             73.0                                                                             89.1                                                                             63.9 a                                                                            6.30 d (J 3.2 Hz)a                                       31   90.1  70.9                                                                             86.6                                                                             62.0 b                                                                            6.13 dd (J 3.0; 0.8 Hz)b                                                                  0.15 h                                       32   90.0                                                                             40.2                                                                             71.0                                                                             86.3                                                                             62.1 b                                                                            6.10 d (J 3.7 Hz)b                                       33   87.8                                                                             40.5                                                                             70.3                                                                             84.9                                                                             61.2 b                                                                            6.23 t (J 3.4 Hz)b                                       34   90.0                                                                             40.2                                                                             71.0                                                                             86.3                                                                             62.1 b                                                                            6.10 d (J 3.1 Hz)                                        35   87.8                                                                             40.5                                                                             70.3                                                                             84.9                                                                             61.2 b                                                                            6.24 t (J 3.3 Hz)                                        62                                         140 (dec.)                         63                                         242 (dec.)                         64                                         227 (dec.)                         65                                         240 (dec.)                         __________________________________________________________________________    Additional data for 1-(2-deoxy-alpha/beta-D-ribofuranosyl)-5-R.sup.2          -uracil compounds                                                             __________________________________________________________________________               Elemental analysis Calculated (found) %                            Example    C               H     N                                            __________________________________________________________________________    25 × 1 H.sub.2 O                                                                   47.54           4.91  8.53                                                    (47.7)          (4.5) 8.5                                          28 × 0.5 H.sub.2 O                                                                 42.63           4.13  7.65                                                    (42.7)          (4.0) (7.7)                                        __________________________________________________________________________     a) CD.sub.3 OD; b) DMSOd.sub.6 c) EtOAc--EtOH 181 d) CHCl.sub.3 --MeOH        8515 e) EtOAc--EtOH 91 f) CHCl.sub.3 --MeOH 71 g) CHCl.sub.3 --MeOH 51 h)     EtOAc--MeOH 91                                                           

Example 36 1-(2,3-Dideoxy-α-D-ribofuranosyl)-5-(2-thienyl)uracil (VSB533)

1-(2,3-Dideoxy-5-O-tert-butyldiphenylsilyl-αD-ribofuranosyl-5-(2-thienyl)uracil(0.15 g) was dissolved in tetrahydrofuran, 1M in tetrabutylammoniumfluoride (3 ml) and stirred at ambient temperature for 1 hour. Thesolvent was evaporated and the product was purified by separation onpreparative thin layer chromatography (silica--1 mm, ethylacetate-methanol 9-1) to give1,(2,3-dideoxy-α-D-ribofuranosyl)-5-(2-thienyl)uracil. TLC (silica,ethyl acetate-methanol 9-1) Rf 0.54.

Example 37 1-(2,3-Dideoxy-β-D-ribofuranosyl)-5-(2-thienyl)uracil (VSB534)

Starting from1-(2,3-Dideoxy-5-O-tert-butyldiphenylsilyl-βD-ribofuranosyl-5-(2-thienyl)uracil(0.35 g) and using the same reaction conditions as described for thecorresponding α-anomer in example 36, the title compound was obtained.TLC (silica, ethyl acetate-methanol 9-1) Rf 0.59.

The starting materials for the α- and β-anomers of1-(2,3-dideoxy-D-ribofuranosyl)-5-(2-ethenyl)uracil (examples 36 and 37respectively) were prepared by the following sequence of reactions a-e.

a) S-γ-tert-Butyldiphenylsilyloxymethyl-γ-butyrolactone (VSB 526)

S-(+)-γ-Trityloxymethyl-γ-butyrolactone (25 g) was mixed with 80% aceticacid (aq, 400 ml) and stirred at 70°-90° C. for 2 hours. The solvent wasevaporated in vacuo and the residue was chromatographed on a column ofsilica, eluted with ethyl acetate-hexane 1-2, to affordS-γ-hydroxymethyl-γ-butyrolactone (VSC 525) as an oil (7.24 g, 90%).This product was dissolved in dry dimethyl formamide (600 ml), imidazole(10.6 g) followed by tert-butyl-diphenylchlorosilane (25 ml, 25.7 g)were added and the solution was stirred at ambient temperature for 4hours and then at 60° C. for another hour. The solvent was evaporated invacuo, the residue was dissolved in ethyl acetate, the solution wasextracted with water and brine, dried (MgSO₄) and the solvent wasevaporated in vacuo. The residue was purified by chromatography on acolumn of silica (ethyl acetate-hexane, 1-4) to yieldS-γ-tert-butyldiphenylsilyloxymethyl-γ-butyrolactone (16.9 g, 77%) m.p.76.5°-77° C.

b) 2,3-Dideoxy-5-O-tert-butyldiphenyl-silyl-D-ribofuranose (VSB 527)

S-γ-tert-Butyldiphenylsilyloxymethyl-γ-butyrolactone (17.1 g) in drydiethyl ether (200 ml) was cooled to -78° C. and stirred whilediisobutylaluminum hydride in hexane (75 ml, 1.1M) was added during15-20 minutes. The stirring was continued for 1 hour at -78° C. afterwhich methanol (35 ml) was added and the reaction solution was allowedto come to room temperature. An aqueous sodium potassium tartratesolution (30%, 150 ml) was added with stirring. The organic phase wasseparated and extracted with the tartrate salt solution (4×75 ml). Thecombined aqueous portions were extracted with diethyl ether (4×75 ml).The combined organic solutions were dried (MgSO₄) and the solvent wasevaporated to give2,3-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranose (16.3 g) as aviscous clear oil.

c) 1-Acetyl-2,3-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranoside(VSB 528)

Acetic anhydride (15 ml) was added dropwise to an ice-cooled solution of2,3-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranose (7.58 g) in drypyridine (25 ml). The stirring was continued at room temperature for 14hours after which the reaction solution was poured onto ice andextracted with diethyl ether. The ether solution was washed with water,followed by a saturated aqueous sodium hydrogencarbonate solution, waterand brine and then dried (MgSO₄). The solvent was evaporated to give1-acetyl-2,3-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranoside as aslightly yellow oil (6.90 g, 89%).

d)1-Dideoxy-5-O-tert-butyldiphenylsilyl-α-D-ribofuranosyl-5-(2-thienyl)uracil(VSB 530) and

e)1-(2,3-Dideoxy-5-O-tert-butyldiphenylsilyl-β-D-ribofuranosyl-5-(2-thienyl)uracil(VSB 529)

5-(2-Thienyl)uracil (0.85 g) was suspended in hexamethyldisilazane (30ml) and chlorotrimethylsilane (0.5 ml) and heated at 90° C. overnighttogether with a small amount of ammoniumsulfate. The solvent wasevaporated in vacuo and the residual bis-trimethylsilyated5-(2-thienyl)uracil was dissolved in dry acetonitrile (10 ml) togetherwith 1-acetyl-2,3-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranoside(1.75 g). The solution was cooled to -35° C. and SnCl₄ (1.14 g, 0.51 ml)in dry acetonitrile (5 ml) was added dropwise. The reaction temperaturewas raised to -15° C. and an excess of ammonia in methanol was added.The solution was allowed to reach room temperature, the solvent wasevaporated in vacuo; the residue was extracted with ethyl acetate,filtered, the solvent was again evaporated in vacuo and the residue wassubjected to chromatography on a column of silica eluted with ethylacetate-hexane 1-9, to give the α- and β-anomers of1-dideoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranosyl-5-(2-thienyl)uracil.

α-anomer: 0.18 g TLC (silica, ethyl acetate-hexane, 1-1) Rf 0.42. ¹³ CNMR (CDCl₃)δ: 26.20 (C3'); 26.98 (CH₃); 32.7 (C2'); 65.80 (C5'); 81.68(C4'); 86.87 (C1')); 109.78 (C5); 125.40, 126.93, 127.2, 133.2(thienyl); 127.76, 127.88, 129.98, 135.64 (phenyl); 133.6 (C6); 149.67(C2); 162 (C4).

β-anomer: 0.38 g, TLC (silica, ethyl acetate-hexane, 1-1) Rf 0.60. ¹³ CNMR (CDCl₃)δ: 26 (C3'); 27 (CH₃); 33 (C2'); 66 (C5'); 82 (C4'); 88.5(C1'); 125, 127, 133 (thienyl); 128, 130, 136 (phenyl); 134 (C6).

Example 38 1-(2,5,6-Trideoxy-α-D-ribo-hexofuranosyl-6-phosphonicacid)-5-(2-thienyl)uracil (VSB 823)

1-(2,5,6-Trideoxy-6-diemthylphosphono-α-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(214 mg) was heated at reflux in hexamethyl-disilazane (5 ml) andacetonitrile for about 15 minutes until all material was dissolved. Thesolvent was evaporated, bromotrimethylsilane (0,2 ml) in acetonitrile (5ml) was added and the solution was stirred at ambient temperature for 3hours. Aqueous ammonia (25%, 5 ml) was added, the solvent was evaporatedand the residue was dissolved in water-dimethyl sulfoxide (about 1 ml).After filtration trifluoroacetic acid (10 drops) and acetone (5 ml) wasadded, the precipitate was collected and washed (decanted) with acetone(3×5 ml), to yield 1-(2,5,6-trideoxy-α-D-ribo-hexofuranosyl-6-phosphonicacid)-5-(2-thienyl)-uracil. TLC (polyethylene imine, Macherey-Nagel,0.2M LiCl, molybdate spray-reagent) Rf 0.15.

Example 39 1-(2,5,6-Trideoxy-β-D-ribo-hexofuranosyl-6-phosphonicacid)-5-(2-thienyl)uracil (VSB 822)

Starting from1-(2,5,6-trideoxy-6-dimethyl-phosphono-β-D-ribo-hexofuranosyl-5-(2-thienyl)uracil(170 mg) and using the same reaction conditions as described for thecorresponding α-anomer (example 38), the title compound was obtained (40mg). TLC (polyethylene imine, Macherey-Nagel, 0.2M LiCl, molybdate sprayreagent) Rf 0.15. ¹³ C NMR (DMSO-d6)δ: 22.70, 25.45 (C5'); 26.96 (C6');39.86 (C2'); 72.06 (C3'); 85.75 (C1'); 88.64, 88.98 (C4'); 108.10 (C5');122.99, 126, 126.83, 134.55 thienyl); 138 (C6'); 149.82 (C2'); 161.62(C4').

The starting materials for the α- and β-anomers of1-(2,5,6-trideoxy-D-ribo-hexofuranosyl-6-phosphonicacid)-5-(2-thienyl)-uracil (examples 38 and 39 respectively) wereprepared by the following reaction sequence (a-h).

a)Methyl-2-deoxy-3-O-p-toluoyl-5-O-tert-butyldiphenylsilyl-D-ribofuranoside

Imidazole (18.9 g) and tert-butyldiphenyl-chlorosilane (37.7 g) wereadded to methyl-2-deoxyribofuranoside (20.3 g) dissolved indimethylformamid (150 ml) and the solution was stirred at ambienttemperature over night. Thin layer chromatography (TLC, silica, ethylacetate-hexane 1-4) shows the reaction productmethyl-2-deoxy-5-O-tert-butyldiphenylsilyl-D-ribofuranoside with Rf 0.2.The solvent was evaporated in vacuo, the residue was dissolved indiethyl ether washed with water (4×50 ml), dried (MgSO₄) and the solventwas evaporated to give a residue (47.1 g). The residue was dissolved inpyridine (200 ml), p-toluoylchloride (21.18 g) was added and thesolution was stirred at ambient temperature for about 1 hour after whichthe solvent was evaporated in vacuo, the residue was taken up in diethylether and washed with water. The solution was dried (MgSO₄) and thesolvent was evaporated in vacuo to give the title compound (50 g). TLC(silica, etyl acetate-hexane 1-4) Rf 0.5. ¹³ C NMR (CDCl₃)δ: 21.77 (CH₃,p-tol.); 26.73, 26.90 (CH₃, tert-but.); 39.41 (C2); 55.41 (OCH₃); 65.02(C5); 75.85 (C3); 84.29 (C4); 105.77 (C1); 127.78, 128.34, 129.17,129.60, 129.77, 134.96, 135.73 (C, phenyl).

b) Methyl-2-deoxy-3-O-p-toluoyl-D-ribofuranoside (VSB 818)

Methyl-2-deoxy-3-O-p-toluoyl-5-O-tert-butyldiphenylsilyl-D-ribofuranoside(50 g) was dissolved in a 1M solution of tetrabutyl-ammonium fluoride intetrahydrofuran (100 ml). Dry sodium hydrogencarbonate (1 eq, 137 mmol)was added and the mixture was stirred at ambient temperature over night;after which the solvent was evaporated, the residue was washed withwater and purified by chromatography on a silica column, eluted withethylacetate-hexane (1-4), followed by ethyl acetate, to give the titlecompound (16.65 g). TLC (silica, ethyl acetate-hexane), 1-4) Rf 0.1. ¹³C NMR (CDCl₃) δ: 21.80 (CH₃, p-tol.); 40.14 (C2); 55.68 (OCH₃); 64.10(C5); 75.97 (C3); 86.35 (C4); 105.84 (C1); 129.24, 129.70, 129.80,129.93 (C, phenyl).

c) Diphenyl(1-methyl-2,5,6-trideoxy-3-O-p-toluoyl-D-ribo-hex-5-enofuranos-6-yl)-phosphonate(VSB 818)

Pyridinium trifluoroacetate (1.89 g) prepared from equimolar amounts ofpyridine and trifluoroacetic acid in diethyl ether) and some molecularsieves (4 Å) were added to a solution ofmethyl-2-deoxy-3-O-p-toluoylfuranoside (5.26 g) in dimethylsulfoxide (40ml). The solution was stirred at ambient temperature for about 30minutes after which dicyclohexylcarbodiimide (12.2 g) was added and thestirring was continued at 60° C. for 3 hours. TLC (silica, ethylacetate-hexane, 1-4) shows a positive reaction with dinitrophenylhydrazin-sulfuric acid spray at Rf 0.1. Methanol (20 ml) was added andstirring was continued at 60° C. for another hour, after which methanolwas evaporated in vacuo, the solution was filtered,diphenyl[(triphenylphosphoranylidene)methyl]phosphonate [9 g; G. H.Jones, E. K. Hamamura, J. Moffat, Tetrahedron Lett. (1968), 5371; J. A.Montgomery, A. G. Laseter, K. Hewson, J. Heterocyclic Chem. 11 (1974)211] was added and the solution was stirred at 70° C. for 3 hours. Aftercooling, diethyl ether (200 ml) was added, the solution was washed withwater (4×100 ml) and the ether solution was evaporated to dryness. Theresidue was purified by chromatography on a column of silica (500 g)eluted with ethyl acetate-hexane 1-4, yielding 4.4 g ofdiphenyl(1-methyl-2,5,6-trideoxy-3-O-p-toluoyl-D-ribo-hex-5-enofuranos-6-yl)phosphonate.¹³ C NMR (CDCl₃) δ: 21.70 (CH₃, p-tol); 37.85 (C2); 56.00 (OCH₃); 77.45(C3); 83.78, 84.24 (C4); 106.52 (C1); 115.33, 119.12 (C5); 152.40,152.52 (C6); 165.97 (CO).

d)Diphenyl(1-methyl-2,5,6-trideoxy-3-O-p-toluoyl-D-ribo-hexofuranos-6-yl)phosphonate(VSB 819)

Diphenyl(1-methyl-2,5,6-trideoxy-3-O-p-toluoyl-D-ribo-hex-5-enofuranos-6-yl)phosphonate(4.46 g) in dry tetrahydrofurane was hydrogenated at 1 bar for 30minutes using Pd/C (5%) as a catalyst. The reaction mixture was filteredthrough a celite pad, the solvent was evaporated and the residue waspurified by chromatography, on silica to give the title compound (3.72g). ¹³ C NMR (CDCl₃) δ: 21.53 (CH₃, p-tol); 21.21, 24.06 (C5); 27.68(C6); 38.95 (C2); 55.27 (OCH₃); 77.52 (C3); 83.68, 84.04 (C4); 105.50(C1); 166.02 (CO).

e)1-(2,5,6)-Trideoxy-3-O-p-toluoyl-6-diphenylphosphono-α-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(VSB 826) and f)1-(2,5,6)-Trideoxy-3-O-p-toluoyl-6-diphenylphosphono-β-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(VSB 820)

5-(2-thienyl)uracil (0.5 g) in dry acetonitrile (15 ml), hexamethyldisilazane (5 ml) and chlorotrimethyl silane (0.5 ml) was heated atreflux for about 30 minutes after which the solvent were evaporated togive 2,4-bis-trimethyl silylated 5-(2-thienyl)uracil. Dry acetonitrilewas added, followed bydiphenyl(1-methyl-2,5,6-trideoxy-3-O-p-toluoyl-D-ribo-hexofuranos-6-yl)phosphonate(VSB 819, 1.65 g) in dry acetonitrile (10 ml) and finallytert-butyl-dimethylsilyltriflate (0.6 ml) under vigorous stirring, andthe solution was stirred at ambient temperature for about 1.5 hours,after which concentrated aqueous ammonia (4 ml) was added). The solventwas evaporated in vacuo and the residue was purified by chromatographyin a column of silica (100 g) eluted with ethyl acetate-hexane, 1-1, togive the α-anomer (0.56 g) and the β-anomer (0.40 g) of1-(2,5,6-trideoxy-3-O-p-toluoyl-6-diphenyl-phosphono-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil.TLC (silica, ethyl acetate-hexane, 1-1) Rf: α 0.15; β 0.20. ¹³ C NMR(CDCl₃) δ, α-anomer: 20.12 (CH₃, p-tol.); 19.68, 22.53 (C5'); 25.40,25.47 (C6'); 36.81 (C2'); 76.50 (C3'); 85.84, 86.16 (C4'); 86.35 (C1');108.22 (C5'); 122.89, 124.23, 125.49 thienyl); 134.03 (C6); β-anomer:21.80 (CH₃, p-tol.); 21.21, 24.08 (C5'); 27.08 (C6'); 37.27 (C2'); 76.48(C3'); 84.12, 84.48 (C4'); 85.70 (C1'); 110.75 (C5'); 124.76, 125.62,127.17 (thienyl); 133.86 (C6).

g)1-(2,5,6)-Trideoxy-6-dimethylphosphono-α-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(VSB 825)

1-(2,5,6-Trideoxy-3-O-p-toluoyl-6-diphenylphosphono-α-D-ribohexofuranosyl-5-(2-thienyl)uracil(444 mg) was dissolved in 0.5M sodium methoxide in methanol (20 ml) andstirred at ambient temperature for 3 hours. The solution was neutralizedwith Dowex 50W×8 (pyridinium⁺), filtered and the solvent was evaporated.Silica and diethyl ether-hexane was added, the solvent was decanted andthe residue was again triturated with ether-hexane (4×). Finally thesilica was eluted with methanol-tetrahydrofuran 1-1 and the solvent wasevaporated in vacuo to give the title compound (234 mg). ¹³ C NMR (CD₃OD) δ: 18.95, 21.80 (C5'); 25.98, 26.08 (C6'); 39.75 (C2'); 52.49 (2POCH₃); 73.32 (C3'); 86.28 (C1'); 88.25, 88.57 (C4'); 109.29 (C5);123.79; 125.10, 126.59, 133.88 (thienyl); 136.59 (C6); 149.92 (C2);161.91 (C4).

h)1-(2,5,6)-Trideoxy-6-dimethylphosphono-β-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(VSB 824)

Starting from1-(2,5,6-trideoxy-3-O-p-toluoyl-6-diphenylphosphono-β-D-ribo-hexofuranosyl)-5-(2-thienyl)uracil(326 mg) and using essentially the same reaction conditions as describedfor the α-anomer, the title compound was obtained. In the work-upprocedure for the β-anomer no silica was included; instead the crudeproduct was dissolved in diethyl ether and by addition of hexane theproduct precipitated (190 mg). ¹³ C NMR (CDCl₃ -DMSO-d6) δ: 18.90, 21.75(C5'); 25.98, 26.08 (C6'); 39.43 (C2'); 52.08 (2 POCH₃); 73.05 (C3');85.38, 85.45, 85.80 (C1', C4'); 109.78 (C5); 123.86, 125.13, 126.52,133.13 (thienyl); 134.57 (C6); 149.38 (C2); 162 (C4);

The precursor 5-substituted pyrimidine compounds of the formula I',##STR15## wherein the radicals R¹ and R² are defined as follows: R¹ :OH, NH₂ ;

R² : ##STR16## wherein X is O, S, N--R⁷, Se;

R⁶ is H, straight or branched C₁₋₁₀ alkyl, F, Cl, Br, I,

X--R⁷, --CH═CH--R⁷, --C≡C--R⁷, CO₂ R⁷, CH₂ X--R⁷ ;

R⁷ is H, straight or branched C₁₋₅ alkyl, phenyl; constitute a furtheraspect of the invention.

The compounds of the formula I' may be prepared by the following generalmethod:

The 2,4-dialkoxy-5-halopyrimidine compound may be reacted with theboronic acid or trialkylstannyl derivative of the heterocycle;alternatively the 2,4-dialkoxy-5-boronic acid pyrimidine or2,4-dialkoxy-5-trialkylstannyl pyrimidine may be reacted with thehalogen derivative of the heterocycle. In all cases the reaction iscatalyzed by a palladium complex and performed in an organic solventsuch as for example tetrahydrofuran or 1,2-dimethoxyethane at atemperature from -20° to 100° C. or at reflux for a period of 5 minutesto 2 days. After completion of the condensation reaction and work-up ofthe reaction mixture the 2,4-dialkoxy groups of the pyrimidine compoundare hydrolyzed by acidic hydrolysis by known methods.

The 5-substituted uracil base or the 5-substituted uridine analogue maybe converted to a 5-substituted cytosine base or cytidine analogue byconventional methods, the principles of which have been described forexample by W. L. Sung (J. Chem. Soc. Chem. Commun. 1981, p. 1089 and J.Organic Chemistry 1982, volyme 47, pages 3623-3628) and by P. Herdewijnet al. (J. Medicinal Chemistry 1985, volyme 28, pages 550-555).

The following examples will further illustrate the precursor compoundsof the invention.

Example 40 5-(5'-Chloro-2'-thienyl)uracil

A 250 ml flask was charged with 3.41 g (0.010 mole) of 2,4-di-tertbutoxy-5-(5'-chloro-2'-thienyl)pyrimidine, 60 ml of methanol and 60 mlof 4M hydrochloric acid and the reaction mixture was stirred at roomtemperature for 30 min. The precipitate crystals were collected byfiltration, washed with methanol and dried giving an almost quantitativeyield of the title compound, mp over 300° C.

Anal. Found C 42.1, H 2.20, N 12.25, S 14.2. Calc. for C₈ H₅ ClN₂ O₂ S(228.6): C 42.02, H 2.20, N 12.25, S 14.02.

The starting material,2,4-di-tert.butoxy-5-(5'-chloro-2'-thienyl)pyrimidine, was prepared asfollows:

A 100 ml flask equipped with condenser, magnetic stirrer and nitrogeninlet was charged with 1.65 g (0.010 mol) of 2-bromo-5-chlorothiophene,0.3 mmol of tetrakis(triphenylphosphine)-palladium(0) and 50 ml1,2-dimethoxyethane. After stirring for 10 min, 2.95 g (0.011 mole) of2,4-di-tert.butoxy-5-pyrimidineboronic acid was added immediatelyfollowed by 20 ml of 1M sodium carbonate solution. The reaction mixturewas refluxed for 4 hours with vigorous stirring under nitrogen. Aftercooling to room temperature the traces of the catalyst were filteredoff, the organic solvent was evaporated under reduced pressure and theresidue was diluted with water and extracted with three portions ofether. The combined etheral phases were washed with water, saturatedsodium chloride solution and dried over magnesium sulphate. The solventwas evaporated and the residue purified by flash-chromatography onsilica gel giving 2.6 g (76%) of2,4ditert.butoxy-5-(5'-chloro-2'-thienyl)pyrimidine mp 82.0°-83.5° C.

Anal. Found C 56.4, H 6.24, N 8.16, S 9.52. Calc. for C₁₆ H₂₁ ClN₂ O₂ S(340.9): C 56.37, H 6.21, N 8.22, S 9.41.

Example 41 5-(3'-furyl)uracil

A 100 ml flash was charged with 1.45 g (5.0 mmole) of2,4-di-tert.butoxy-5-(3'-furyl)pyrimidine dissolved in 25 ml of methanoland 25 ml of 5M hydrochloric acid and the mixture was stirred at roomtemperature for 30 min. The precipitated crystals were collected byfiltration, washed with methanol and dried giving the title compound inalmost quantitative yield, melting with decomposition above 250° C.

Anal. C 54.1, H 3.34, N 15.5, O 27.2. Calc. for C₈ H₆ N₂ O₃ (178.1): C53.9, H 3.39, N 15.7, O 26.9.

The starting material 2,4-di-tert.butoxy-5-(3'-furyl)pyrimidine wasprepared as follows:

A 250 ml flask equipped with condenser, magnetic stirrer and nitrogeninlet was charged with 7.3 g (0.024 mole) of5-bromo-2,4-di-tert-butoxypyrimidine, 0.75 mmol oftetrakis(triphenylphosphine)palladium (0) and 80 ml of1,2-dimethoxyethane. After stirring for 10 min 3.0 g (0.027 mole) of3-furanboronic acid was added, immediately followed by 60 ml of 1Msodium carbonate solution. The reaction mixture was refluxed for 4 hourswith vigorous stirring under nitrogen. After cooling to roomtemperature, the traces of catalyst were filtered off, the organicsolvent was evaporated under reduced pressure and the residue dilutedwith water and extracted with three 50 ml portions of ether. Thecombined etheral phases were washed with water, saturated sodiumchloride solution and dried over magnesium sulphate. The ether wasevaporated and the residue was purified by flash chromatography usinghexane-ethyl acetate (4:1) as eluent, yielding 4.1 g (59%) of the titlecompound as an oil.

Anal. Found C 66.5, H 7.68, N 9.64, O 17.0. Calc. for C₁₆ H₂₂ N₂ O₃(290.4) C 66.2, H 7.64, N 9.65, O 16.5.

Example 42 5-[2'-(N-methyl)pyrrolyl]uracil

3.0 g (9.9 mmole) of2,4-di-tert.butoxy-5-[2'-(N-methyl)pyrrolyl]-pyrimidine was stirred with40 ml of methanol and 40 ml of 5M hydrochloric acid for 30 min. Theprecipitated crystals were collected by filtration, washed with methanoland water and dried, yielding 1.5 g (79%) of the title compound meltingwith decomposition over 250° C.

Anal. Found C 56.0, H 4.70, N 22.00. Calc. for C₉ H₉ H₃ O₂ (191.2): C56.5, H 4.47, N 22.0.

The starting material2,4-di-tert.butoxy-5-[2'-(N-methyl)pyrrolyl]pyrimidine was prepared asfollows:

A 250 ml flash equipped with condenser, magnetic stirrer and nitrogeninlet was charged with 9.0 g (29.7 mmole)2,4-di-tert-butoxy-5-bromopyrimidine. 1.05 g (1.50 mmole) of PdCl₂ [P(C₆H₅)₃ ]₂ and 8.0 g (32.7 mmole) of N-methyl-2-trimethylstannylpyrrole in80 ml of anhydrous tetrahydrofuran and refluxed for 20 hours. Aftercooling the reaction mixture, it was diluted with 200 ml of ether andwashed twice with 50 ml of water. After drying with magnesium sulphateand evaporating the solvent, the compound was purified by chromatographyusing "silicagel 60" and a mixture of pentane-ether (9:1) as eluent,yielding 3.5 g (39%) of the title compound, mp 113°-114° C.

Anal. Found C 67.0, H 8.37, N 13.7. Calc. for C₁₇ H₂₅ N₃ O₂ (303.4) C67.3, H 8.30, N 13.8.

Analogous to example 40, table 5 gives some further examples ofpreparations from 2,4-di-tert.-butoxy-5-pyrimidine boronic acid and abromo substituted heterocyclic compound. Their characteristics are givenin table 6.

                  TABLE 5                                                         ______________________________________                                        Examples of 5-R.sup.2 -uracil compounds                                                      Intermediate                                                                  2,4-di-tert-                                                                  butoxy-5-(R.sup.2)                                                            pyrimidine  5-R.sup.2 -uracil                                  Example                                                                              R.sup.2       yield %  mp °C.                                                                        yield                                    ______________________________________                                        40     2-(5-chloro)thienyl                                                                         76         82-83.5                                                                             86                                      43     2-(5-methyl)thienyl                                                                         50       65-68   93                                      44     2-(5-hexyl)thienyl                                                                          52       oil     90                                      45     2-furyl       49       87-88  100                                      46     2-thiazolyl   49       102-103                                                                              100                                      47     5-thiazolyl   62       68-69  100                                      48     2-pyridyl     60       128-129                                                                              100                                      49     3-pyridyl     69       88-89  100                                      50     4-pyridyl     70       92-93  100                                      51     2-methoxyphenyl                                                                             35         90-91.5                                                                             90                                      52     3-methoxyphenyl                                                                             65       93-94   90                                      53     4-methoxyphenyl                                                                             41       92-94   90                                      54     2,5-dimethoxyphenyl                                                                         47       91-93  100                                      66     2-trans-tioften                                                                             57       108-110                                         67     2-cis-tioften 60       108-110                                         68     3-trans-tioften                                                                             55       105-107                                         69     3-cis-tioften 27       88-90                                           ______________________________________                                    

                                      TABLE 6                                     __________________________________________________________________________    .sup.1 H NMR chemical shifts (ppm, in DMSO-d.sub.6) for 5-substituted         uracil compounds                                                              Example                                                                            NH NH H6 H2'                                                                              H3'                                                                              H4'                                                                              H5'                                                                              H6'                                                                              CH.sub.3                                                                         OCH.sub.3                                                                         OCH.sub.3                                                                         CH.sub.2                              __________________________________________________________________________    40   11.5  8.07  7,03                                                                             7.33                                                                             -- --                                                  43   9.3   7.83  7.22                                                                             6.71                                                                             -- -- 2.51       2.73 a                                44   11.39                                                                            11.20                                                                            7.83                                                                             -- 7.24                                                                             6.73                                                                             -- --                                                  45   11.40                                                                            11.20                                                                            7.71                                                                             -- 6.83                                                                             6.57                                                                             7.62                                                   46   11.85                                                                            11.78                                                                            8.55                                                                             -- -- 7.92                                                                             7.71                                                                             --                                                  47   11.56                                                                            11.61                                                                            8.22                                                                             9.11                                                                             -- 8.39                                                                             -- --                                                  48   11.90                                                                            11.74                                                                            8.49                                                                             -- 8.67                                                                             8.20                                                                             7.58                                                                             8.33                                                49   11.69                                                                            11.58                                                                            8.13                                                                             9.12                                                                             -- 8.70                                                                             7.98                                                                             7.76                                                50   12.00                                                                            11.66                                                                            8.43                                                                             8.77                                                                             8.37                                                                             -- 8.37                                                                             8.77                                                51   11.17                                                                            10.97                                                                            7.38  7.03                                                                             7.30                                                                             6.93                                                                             7.20  3.73                                          52   11.20 7.65                                                                             7.13                                                                             -- 6.86                                                                             7.26                                                                             7.13                                                53   11.19                                                                            11.02                                                                            7.53                                                                             7.48                                                                             6.94                                                                             -- 6.90                                                                             4.75  3.76                                          54   11.17                                                                            10.95                                                                            7.39                                                                             -- 6.95                                                                             6.85                                                                             -- 6.85  3.70                                                                              3.67                                      __________________________________________________________________________     a) Additional bands at δ 1.64; 1.32; 0.88 with the relative             intensities 2:6:3                                                        

Biological Tests Test I Effect of compounds of the formula I on HIV inH9 cells Materials and methods: HIV infection of H9 cells

H9 cells, 10⁵ cells per well on a 24 well plate, suspended in 2 mlRPMI-medium containing 10% fetal calf serum, 100 μg/ml penicillin, 10μg/ml streptomycin sulfate and 2 μg/ml polybrene are exposed to HIV(HTLV-III_(B)) and different concentrations of the test compounds. Theplates are incubated at 37° C. in 5% CO₂ for 6-7 days. The contents ineach well is then homogenized with a pipette and transferred to acentrifuge tube. After centrifugation for 10 min at 1500 rpm thesupernatant is removed and the cell pellet is analyzed by fixing inmethanol on glass plates. Human HIV positive serum diluted 1:80 or 1:160is added and incubated for 30 min at 37° C. The plate is then washedwith phosphate-buffered saline (PBS) containing Ca²⁺ and Mg²⁺. Sheepantihuman conjugate (FITC) is added and after a new incubation the plateis again washed with PBS. Contrast staining is done with Evans blue andafter drying the frequency of HIV antigen containing cells is determinedin a microscope. The test result is shown in Table 7.

                  TABLE 7                                                         ______________________________________                                        Concentration (μM) for 50% inhibition (IC.sub.50) of human                 immuno deficiency virus multiplication in cell culture                        1-(2'-deoxy-α/β-D-ribofuranosyl)-5-R.sup.2 -uracil                 α/β                                                                        R.sup.2        Code     IC.sub.50 M                                   ______________________________________                                        α 2-thienyl      VSA 134  0.05-10                                       α 2-selenienyl   VSA 188  2-20                                          α 3-selenienyl   VSA 996  3-100                                         α 2-furyl        VSB 007  <10                                           α 2-(5-methylthienyl)                                                                          VSB 515  10->10                                        β  3-selenienyl   VSA 992   5->10                                        β  2-thienyl      VSA 189  10->10                                        β  2-furyl        VSB 008  10->10                                        ______________________________________                                    

Table 7 shows that the tested compounds are active inhibitors of HIVvirus multiplication.

Test II Cellular toxicity

H9 cells, 2×10⁷ cells per plate, are incubated in RPMI-1640 mediumcontaining 10% fetal calf serum, 70 mg/l penicillin, 100 mg/lstreptomycin and 10 mM hepes, in absence or presence of test compounds.The number of cells per plate is determined after 48 h. Cells incubatedin the absence of test compounds then underwent two cell divisioncycles.

F5000 cells, which are human embryo cells, 1×10⁵ cells per plate, areincubated in Eagle's minimal essential medium, supplemented with Earle'ssalts, non-essential amino acids, 10% fetal calf serum, 10 mM hepes, 70mg/l penicillin and 100 mg/l streptomycin, in absence or presence oftest compounds. The number of cells per plate is determined after 48 h.Cells incubated in the absence of test compounds underwent one celldivision cycle. The results are given as % inhibition of cellmultiplication when the concentration of the compound is 100μM or 250μM.The test results are given in table 8.

                  TABLE 8                                                         ______________________________________                                        Cellular toxicity on H9 and F5000 cells                                       1-(2'-deoxy-α/β-D-ribofuranosyl)-5-R.sup.2 -uracil                                     % inhibition                                                                  (concentration μM)                                     α/β                                                                      R.sup.2       Code      H9     F5000                                    ______________________________________                                        α                                                                             2-thienyl     VSA 134   35(250)                                                                              0-35(100)                                α                                                                             2-selenienyl  VSA 188   40(200)                                                                              15(200)                                  α                                                                             3-selenienyl  VSA 996   65(200)                                                                              30(200)                                  α                                                                             2-furyl       VSB 007                                                   α                                                                             2-(5-methyl)thienyl                                                                         VSB 515                                                   β                                                                              3-selenienyl  VSA 992   40(200)                                                                              0(200)                                   β                                                                              2-thienyl     VSA 189   35(200)                                                                              10(100)                                  β                                                                              2-furyl       VSB 008          0(200)                                   ______________________________________                                    

Table 8 shows that the concentrations at which the compounds exhibittoxicities exceed the concentrations needed for 50% inhibition of HIVmultiplication as given in table 7.

Test III Inhibition of reverse transcriptases and DNA polymerases bytriphosphates of compounds of the invention

The 5'-triphosphates were synthesized essentially as described(Yoshikawa, M, Kato T, Takenishi T, Bull. Chem. Soc. (Japan),42,3505-3508, 1969; Ludwig, J., Acta Biochim. Biophys. Acad. Sci. Hung.16, 131-133, 1981; Ruth, J. L., Cheng, Y. C., Mol. Pharmacol. 20, 4151981.) The HIV-RT was obtained as described by Hansen et al (Hansen J,Schulze T and Moelling K, J. Biol. Chem. 262, 12393-12396, 1987) fromcultures of Escherichia coli expressing the cloned HIV-pol gene. TheHBV-DNAP was prepared from virus obtained from human serum, essentiallyas described by Nordenfelt et al (1987) [Nordenfelt E, Lofgren B,Chattopadhyaya J., Oberg B, J. Med Virol. 22, 231-236, 1987]. The HSV-2DNAP and cellular DNAPα preparation and reaction conditions have beendescribed by Larsson et al [Larsson A, Sundqvist A, Parnerud A-M, 1986,Mol. Pharmacol. 29, 614-621]. In reactions using HIV-RT, the enzyme wasincubated with the template (rA)_(n) (dT)₁₂₋₁₈ and differentconcentrations of inhibitor and substrate (dTTP) as described by Vranget al 1987, (Vrang L., Bazin H., Remaud G., Chattopadhyaya J. and ObergB., Antiviral Res. 7, 139-145, 1987). The hepatitis B virus enzymeactivity was determined with a virus preparation solubilized by non-idetP40, and endogenous nucleic acid as template, as described by Nordenfeltet al (vide supra)

                  TABLE 9                                                         ______________________________________                                        Concentration (uM) for 50% inhibition (IC.sub.50) of enzymes                  by triphosphates of some compounds of the invention                                       HIV     HBV       HSV-2                                           Compound    RT.sup.1)                                                                             DNAP.sup.2)                                                                             DNAP.sup.3)                                                                          DNAPα.sup.4)                       ______________________________________                                        1-(2-Deoxy-beta-D-                                                                        0.015   0.11      0.06    1,6                                     ribofuranosyl)-5-                                                             (2-thienyl)uracil-                                                            5'-triphosphate                                                               1-(2-Deoxy-alpha-D-                                                                       2.0     18.0      11.0   80,0                                     ribofuranosyl)-5-                                                             (2-thienyl)uracil-                                                            5'-triphosphate                                                               ______________________________________                                         .sup.1) Human immuno deficiency virus reverse transcriptase                   .sup.2) Hepatit B virus DNA polymerase                                        .sup.3) Herpes simplex virus type 2 DNA polymerase                            .sup.4) DNA polymerase alpha.                                            

I claim:
 1. A compound of the formula ##STR17## wherein R¹ and R² aredefined as follows R¹ is OH, NH₂ ;R² is ##STR18## wherein X is O, S,N--R⁷, Se; R⁶ is H, straight or branched C₁₋₁₀ alkyl, F, Cl, Br, I,X--R⁷, --CH═CH--R⁷, --C═C--R⁷, CO₂ R⁷, CH₂ X--R⁷ ; R⁷ is H, straight orbranched C₁₋₁₅ alkyl, phenyl; with the provisos that when R¹ is OH, R²is not ##STR19## when R⁶ is H, straight or branched C₁₋₁₀ alkyl, orX--R⁷ when X is O and R⁷ is H, or straight or branched C₁₋₁₅ alkyl; andwhen R¹ is NH₂, R² is not ##STR20##
 2. A compound of the formula:##STR21## wherein R¹ and R² are defined as follows: R¹ is OH, NH₂ ;R² is2-thienyl, 2-seleniynyl, 2-furyl, 2-thiazolyl, 2-(1-methyl)pyrrolyl ormethoxyphenyl, with the proviso that when R¹ is OH, R² is not 2-thienyl,2-seleniynyl or methoxyphenyl.